Epstein-Barr Virus: Activation of WNT

Epstein-Barr 病毒：WNT 激活

• 批准号: 7105558
• 负责人: NANCY JOAN RAAB-TRAUB
• 金额: $ 31.72万
• 依托单位: UNIV OF NORTH CAROLINA CHAPEL HILL
• 依托单位国家: 美国
• 财政年份: 2004
• 资助国家: 美国
• 起止时间: 2004-08-11 至 2009-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7105558
• 关键词: Epstein Barr virus; SCID mouse; athymic mouse; cadherins; cell growth regulation; cell line; clinical research; gel mobility shift assay; human tissue; immunocytochemistry; immunogenetics; protein localization; protooncogene; viral carcinogenesis; virus cytopathogenic effect; virus protein; virus related neoplasm /cancer

DESCRIPTION (provided by applicant): The goal of this proposal is to characterize the effects of Epstein Barr Virus (EBV) on the Writ/Wingless signaling pathway which is a critical regulator of cell growth and differentiation during development and is frequently affected during cancer development. In this application, we will determine if this pathway is activated infected cell lines and in EBV-associated cancers, identify the viral proteins that affect regulation of this pathway, determine the mechanisms that underlie this activation, and identify the cellular targets that are affected by this activation. Specifically we will: 1) Characterize beta-catenin levels, localization, and activity in EBV positive and negative cell lines and in EBV-associated and AIDS-related cancers using cell fractionation and immunoblotting, electrophoretic mobility shift assays (EMSA) on beta-catenin responsive elements, reporter assays, and immunohistochemistry to identify specific phosphorylated protein components of the pathway and determine intracellular localization of beta-catenin. 2) Identify the effects of LMP 1 on expression and localization of components of writ/wingless using protein and gene array analyses. 3) Determine the effects of LMP2 on Akt activation, inactivation of glycogen synthase kinase beta, B-catenin localization, and wnt-regulated gene expression. Compare and contrast LMP2 with mutated forms of LMP2 and with constitutively activated forms of PI3-kinase and Akt. Determine the contribution of activation of this pathway on LMP2-mediated effects on cellular growth using fibroblast and epithelial cell transformation assays and perform genomic expression array analysis on cell lines expressing LMP2, mutant forms of LMP2, or activated components of wnt signaling to determine effects on gene expression. 4) Analyze LEF/TCF regulated gene expression in cell lines expressing EBV latent proteins. Identify components of the transcription complex and determine how EBV proteins affect this complex. The proposed studies represent the first studies on a completely new pathway likely affected in EBV-associated cancers. These studies will determine the contribution of this pathway in the development of EBV-associated cancers and identify the specific effects of EBV proteins on the regulation of wnt/wingless signaling. A detailed understanding of the activation of this pathway may provide new opportunities for treatment of EBV-associated cancers and also of similar cancers considered EBV-negative, that develop in the immunosuppressed.

描述（由申请人提供）：本申请的目标是表征eb病毒（EBV）对Writ/Wingless信号通路的影响，该信号通路是发育过程中细胞生长和分化的关键调节因子，在癌症发展过程中经常受到影响。在本应用中，我们将确定这一途径是否在感染细胞系和ebv相关癌症中被激活，确定影响这一途径调控的病毒蛋白，确定这种激活的机制，并确定受这种激活影响的细胞靶标。具体来说，我们将：1)利用细胞分离和免疫印迹、β -连环蛋白反应元件的电泳迁移位移测定（EMSA）、报告者测定和免疫组织化学来鉴定该途径的特定磷酸化蛋白组分，并确定β -连环蛋白的细胞内定位，表征EBV阳性和阴性细胞系以及EBV相关和艾滋病相关癌症中β -连环蛋白的水平、定位和活性。2)利用蛋白质和基因阵列分析确定lmp1对writ/wingless组分表达和定位的影响。3)确定LMP2对Akt活化、糖原合成酶激酶β失活、B-catenin定位和wnt调控基因表达的影响。将LMP2与突变形式的LMP2以及组成型活化形式的pi3激酶和Akt进行比较。利用成纤维细胞和上皮细胞转化试验确定该途径的激活对LMP2介导的细胞生长效应的贡献，并对表达LMP2、LMP2突变形式或wnt信号激活成分的细胞系进行基因组表达阵列分析，以确定对基因表达的影响。4)分析LEF/TCF在EBV潜伏蛋白表达细胞系中的调控基因表达。鉴定转录复合体的组分，并确定EBV蛋白如何影响该复合体。拟议的研究是对ebv相关癌症可能受影响的全新途径的首次研究。这些研究将确定这一途径在EBV相关癌症发展中的作用，并确定EBV蛋白对wnt/无翅信号传导调节的特定作用。对这一途径激活的详细了解可能为治疗ebv相关癌症以及被认为是ebv阴性的类似癌症提供新的机会，这些癌症在免疫抑制中发展。